Antibodies against recombinant heat shock protein 65 of Tropheryma whipplei in patients with and without Whipple's disease

Tropheryma whipplei is the causative agent of Whipple's disease (WD). a chr onic, life-threatening infection. Laboratory diagnosis is mainly based on P CR and histopathological analysis in duodenal biopsies and other specimens requiring invasive procedures. We have examined the presence of antibodies to recombinant heat shock protein (Hsp65) of T. whipplei in patients with W hipple's disease as well as in control subjects by Western blot and enzyme- linked immunosorbent assay (ELISA). A recombinant plasmid carrying the enti re T. whipplei hsp65 gene was constructed, and the expression yielded a 65- kDa histidine-tagged protein. Among four patients with Whipple's disease, t wo showed an IgG- and one an IgA-response, respectively, when analyzed by W estern blotting, whereas from 10 patients without Whipple's disease, only t wo patients showed a positive IgG-response. The differences between the ser a from patients and controls were thus not significant, Successful purifica tion of the protein was achieved by Ni-NTA affinity chromatography. Quantit ative analysis of serum antibodies by ELISA demonstrated that antibody leve ls in the sera of 14 patients were not significantly higher than in those o f 89 control subjects. The established ELISA test is not useful to clinical diagnostics. (C) 2001 Elsevier Science B.V. All rights reserved.